Myofascial pain syndrome (MPS) is a condition associated with regional pain and muscle tenderness characterized by the presence of hypersensitive nodules, also called myofascial trigger points (MTrPs). MPS affects up to 95% of people with chronic pain disorders and is a common finding in patients in specialty pain management centers. However, the etiology of myofascial pain and the pathophysiology of MTrPs are unclear. MTrPs are palpable, stiff nodules in a taut band of skeletal muscle - a characteristic finding in MPS. Although digital palpation is considered the gold standard for diagnosis of MPS, it does not 1) provide an objective, reliable and sensitive method of diagnosis and measurement of treatment efficacy; 2) provide quantitative comparisons of the tissue properties before and after treatment; 3) objectively differentiate among active MTrPs, latent MTrPs, and palpably normal tissue; 4) objectively discriminate between superficial and deep MTrPs; and 5) permit objective study of the natural history of MTrPs. To address these limitations, our team is utilizing novel applications of diagnostic ultrasound imaging techniques, such as grayscale (2D Ultrasound), vibration sonoelastography, and Doppler, to image the MTrP and its surrounding muscle as well as the local microvasculature and blood flow properties of muscle harboring MTrPs. Through the use of these techniques, our team sought to better understand the physical properties and vascular environment of active MTrPs, latent MTrPs, and palpably normal tissue. Our objectives also included developing new, efficient, and easy to administer elastography techniques to provide quantitative characterization of viscoelastic properties of skeletal muscle in an outpatient clinical environment using off-the-shelf office-based equipment. The sonoelastography studies showed that active MTrPs had larger areas compared to latent MTrPs and palpably normal tissue and therefore MTrPs may be classified by area. Doppler spectral waveform data indicated that blood vessels near these active MTrP sites had a significantly higher pulsatility index compared to normal sites, a finding that may be useful in evaluating the history of MPS. Vibration sonoelastography demonstrated that MTrPs in upper trapezius had significantly higher shear modulus (measure of rigidity) and active MTrPs had significantly higher shear wave speeds (related to viscoelastic properties) when compared to palpably normal tissue. These results indicate that MTrPs and their surrounding tissue have increased tissue heterogeneity. Our research has yielded a better understanding of the pathophysiology of myofascial pain and has implications for clinical care. The sonoelastography techniques can be easily translated to the clinical setting using off the shelf office-based equipment to quantitatively characterize biomechanical properties of skeletal muscle. Furthermore, it can overcome the subjectivity of palpation and physical examination, and could lead to an objective clinical outcome measure. We believe that longitudinal monitoring of biomechanical properties of both MTrPs and the surrounding muscle tissue over time and before and after treatment can provide a better mechanistic understanding of the changes that occur in the environment of the upper trapezius following treatment for active MTrPs, such as dry needling therapy.